Work on the role of insulin-like growth factors (IGFs) in pediatric sarcomas is continuing to focus on potential therapeutic approaches as well as further defining the downstream signaling events of IGF action. We have recently received both humanized moAbs directed against the IGFIR as well as small molecule inhibitors against the IGFIR kinase and have just begun to analyze these reagents to determine whether they specifically inhibit metastatic behavior in several mouse models of metastasis in pediatric sarcomas. We ultimately plan to combine this treatment with inhibitors of mTOR. We have continued to study the role of mTOR signaling in pediatric sarcomas. We have linked ezrin expression and metastatic potential (see below) to mTOR activation and have reported that mTOR blockade using rapamysin and its analog, CCI 779, in rhabdomyosarcomas (RMS) and osteosarcomas inhibits both primary tumor growth and metastatic tumor growth and inhibits mTOR signaling in these tumors. We have recently demonstrated that one mechanism of growth inhibition by rapamycin appears to be via a HIF1alpha, VEGF inhibitory signal. We have also begun to analyze the potential additive or synergistic effects of combining standard cytotoxic chemotherapy with mTOR inhibition in vitro in RMS cell lines and have seen at least additive effects. We are currently beginning to examine this combination in xenograft models. We have expanded our proteomic analysis of stage III RMS tumor samples from patients treated on COG protocols and expanded analysis continues to suggest that activation of mTOR signaling pathways in these tumor samples is a poor prognostic factor. This data has been presented at ASCO and is currently being prepared for publication. We plan to continue to expand these observations with larger patient numbers. Our work on the role of ezrin in Ewing's sarcomas (ES) has now shown that ezrin is very highly expressed in almost all tumors, and its high expression is associated with activation of anti-apoptotic signaling cascades. We have also demonstrated that blockade of ezrin inhibits metastatic behavior in ES xenografts in a manner analogous to OS models. Current work is aimed at determining whether ezrin is linked to mTOR signaling in these tumors. We have also begun to study the role of chemokine signaling in metastatic behavior of OS, and now demonstrated that blocking CXCR5 in OS xenografts blocks metatatic behavior. Ongoing work is aimed at better understanding this interaction to determine whether such approaches may warrent clinical studies.